1. Technical Field
The present invention relates to a macroarray spotting pin for spotting a spotting solution containing biopolymers on a water-absorbing support, such as a nylon membrane, during the macroarray manufacturing process.
2. Background Art
Macroarrays are conventionally manufactured by spotting multiple kinds of spotting solutions containing biopolymers such as DNA, RNA, and proteins on a support, such as a nylon membrane. FIG. 13 illustrates the principle of manufacture of a macroarray. A microplate 132 houses multiple kinds of spotting solutions to be spotted, including a DNA solution 131. The support for the macroarray is comprised of a nylon membrane 134. The DNA solution 131 is carried by a spotting pin 133 and then spotted on the nylon membrane 134, and this process is repeated, thereby producing a plurality of macroarrays 135 on which the multiple kinds of DNA solutions are spotted. Various types of spotting pins for the manufacture of macroarrays have been developed. Examples include a split-type pin capable of sequential spotting based on the capillary action similar to that which occurs in the fountain-pen tip, and a solid-type pin in which a spotting solution is caused to adhere to the pin tip before each stamping.
In order for the results obtained from the macroarray to be reliable, it is necessary to accurately grasp how much of the spotting solution containing biopolymers such as DNA, RNA and proteins is fixed at each spot on the macroarray. It is difficult, however, to quantitatively spot with the solid-type pins. While the split-type pins are advantageous in that they do not require the solution to be adhered to the tip of the pins before each spotting and that they are resistant to drying, for example, it is still difficult to sequentially spot equal amounts.
It is an object of the invention to provide a spotting pin capable of spotting equal amounts of multiple kinds of spotting solutions containing biopolymers on a water-absorbing support used in biological experiments in a stable and sequential manner.
The above object is achieved by a spotting pin according to the invention which is capable of carrying a solution based on the capillary phenomena and which can have its tip split.
In one aspect, the invention provides a spotting pin for spotting a solution on a water-absorbing support, comprising:
a first member comprising a solution holding portion opening into a front and back surface of the tip of the first member which comes into contact with a support, the first member holding a predetermined amount of solution in the solution holding portion based on a capillary action, the first member further comprising a slide guide portion;
a second member comprising a solution supply portion opening into an end of the second member opposite the opening on the back surface of the solution holding portion, the solution supply portion holding the solution based on a capillary action, wherein the second member slides along the slide guide portion of the first member; and
a biasing member for urging the second member against the first member such that the solution supply portion of the second member comes into contact with the solution holding portion of the first member.
By bringing the solution supply portion of the second member into contact and communication with the solution holding portion of the first member, the solution in the solution supply portion can be filled into the solution holding portion of the first member by a capillary action. Then, the second member is caused to slide relative to the first member against the force of the biasing member, in order to separate the solution holding portion of the first member and the solution supply portion of the second member. As a result, a predetermined amount of the solution is carried in the through-hole of the first member due to a capillary action. Next, the tip of the first member is brought into contact with the absorptive support, so that the predetermined amount of solution carried by the solution holding portion of the first member is absorbed into the absorptive support, forming a spot. Thereafter, the solution supply portion of the second member is brought into contact with the solution holding portion of the first member by the force of the biasing member. Consequently, the solution holding portion of the first member that has been empty can be re-filled with the solution from the solution supply portion due to a capillary action. By repeating this sequence, equal amounts of the solution can be sequentially spotted on the absorptive support.
The second member may comprise a body and a branch portion extending from the body in a direction opposite the tip of the first member. In this case, the branch portion acts as a mount via which the spotting pin can be fixed to the pin head of spotting equipment. The first member is driven relative to the second member by a pin or the like protruding from the pin head of the spotting equipment.
The second member may comprise a body and a branch portion extending from the body in a direction of the tip of the first member, and the tip of the branch portion may protrude beyond the tip of the first member when the solution supply portion of the second member is in contact with the solution holding portion of the first member. In this case, the branch portion comes into contact with the support and thus functions as a stopper for separating the solution holding portion of the first member from the solution supply portion of the second member. The spotting pin is fixed to the spotting equipment by having the rear end of the first member fixed to the pin head.
By providing the second member with a large-sized solution reservoir communicated with the solution supply portion, a large amount of a biopolymer solution can be supplied to the spotting pin, so that more spots can be created at once by a single charging of the solution. In this case, a line connecting the tip of the first member and the center of the solution reservoir may be either parallel or non-parallel to the sliding direction of the second member.
Preferably, the periphery of the tip surface of the first member is cut in order to reduce the area of contact with the support. It is also preferable that the periphery of the tip of the second member opposite the back surface of the tip of the first member be cut in order to reduce the area of contact with the back surface of the tip of the first member. By thus cutting the periphery of the tip of the first member and that of the solution supply end of the second member, the movement of the solution by a capillary action can be facilitated, making it possible to create solution spots of the same shape on the support such as a highly water-absorbing nylon membrane in a stable and sequential manner.
The biasing member may be a compression spring disposed between the inner wall of the rear end of the first member and the second member. The compression spring acts to push the second member in the direction of the tip of the first member.
The first and second members may be made of austenitic stainless steel. By using austenitic stainless steel as the material for the spotting pin, the strength and acid and chemical resistance can be improved.
To realize a smooth movement of the spotting pin and extend its life, the sliding portions of the first and second members are preferably diamond-coated.
In another aspect, the invention provides a spotting pin for spotting a solution on a water-absorbing support, comprising:
a first member comprising a plurality of solution holding portions each having an opening on a front and back surface of the tip of the first member which comes into contact with the support, and a slide guide portion, each solution holding portion holding a predetermined amount of the solution based on a capillary action;
a second member comprising a plurality of solution supply portions each having an opening on an end of the second member opposite the opening on the back surface of the solution holding portion and holding the solution based on a capillary action, wherein the second member slides along the slide guide portion of the first member; and
a biasing member for urging the second member against the first member such that the multiple solution supply portions of the second member come into contact with the multiple solution holding portions of the first member.
This spotting pin is an application of the principle of the spotting pins described above, and it comprises a plurality of solution supply portions and pin tips that are connected to one another. This embodiment allows multiple spots to be simultaneously formed on the water-absorbing support. By making the first and second members with plastics, a disposable spotting pin can be provided at reduced costs. Further, contamination of the solution, which is potentially problematic for recycling purposes, can be avoided.
The spotting pin according to the invention can be used for spotting any kind of biopolymers, such as DNA, RNA, proteins, and mixtures thereof. As the water-absorbing support, film-like supports in general with water-absorbing properties for macroarray purposes, such as nylon membranes, can be used.